Normal tissue develops, and is maintained by, processes of cell division and cell death. In many diseases, such as cancer, diabetes mellitus Type I, and autoimmune disease, the normal balance between cell division and cell death is disrupted, causing either a rapid growth of unwanted and potentially dangerous cells, and/or a loss of cells essential to maintaining the functions of tissue. Inappropriate cell division or cell death can result in serious life-threatening diseases. Diseases associated with increased cell division include cancer and atherosclerosis. Diseases resulting from increased cell death include AIDS, neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, retinitis pigmentosa), aplastic anemia, atherosclerosis (e.g., myocardial infarction, stroke, reperfusion injury), and toxin-induced liver disease.
The immune system, a complex organization of cells, tissues and organs, serves to protect us from potential harm. Extraordinary advances in our understanding of the immune system have been made in the last hundred years, especially since the discovery of the T cell and B cell [3-5]. Native T-cells require two signals for activation. These are recognition of antigens in Major Histocompatibility Complex-encoded (MHC) molecules [3], and a co-stimulation signal [6-10] provided by the B7/CD28 family members or other co-stimulatory molecules such as Fas (CD95) [11]. Previously activated T cells can be reactivated by co-stimulation alone [12, 13]. In the absence of activation, T-cells disregard the tissue. If a T cell is activated the consequences can be: 1) destruction of the damaged cells or 2) repair of damaged cells by promoting regeneration either directly or indirectly.
A natural question is: why doesn't the immune system destroy tumor cells as they arise? In fact, there is substantial evidence that the immune system does play an extensive role in suppressing cancer. For example, it is known that people on immune-suppressive therapy have a much higher cancer rate as do people with AIDS [14] and the very young and very old. However, it is also clear that the ability of the immune system to control cancer is not perfect.
Researchers have for many years tried to stimulate the immune system as a therapeutic strategy against cancer [15, 16]. These attempts have generally been ineffective, although there have been some recent successes [17]. There are many reasons for this variability. These include: 1) the inability to activate T cells that can destroy the tumor due to the absence of signal one (lack of recognition of the appropriate tumor antigen); 2) the presence of a signal two that results in the production of the wrong cytokines by T cells which may lead to the growth of tumors, or 3) the failure of activated T cells to kill cancerous cells [18].
Anti-cancer agents may work to promote the death of tumor cells in multiple ways. First, these agents may work by direct cytolysis requiring active participation of the tumor cell in the death process [25]. Second, chemotherapeutics may promote the ability of the tumor cell to be recognized by cells of the immune system and to be killed by immune-directed cell death [26]. Third, these agents may work to “rewire” the death inducing receptor/ligand pairs which include Fas and FasL [23, 25, 26]. The second and third possibilities are not mutually exclusive and may work in concert to result in tumor cell death.
It has been discovered that uncoupling protein (“UCP”) is normally present in the plasma membrane of rapidly dividing cells, but it is not typically found on the plasma membrane of growth-arrested or chemotherapy-resistant tumor cells. UCP, which is also present in the mitochondria, can regulate cell division by manipulating the manner in which the cell processes and stores energy. It was found that the UCP in the plasma membrane plays an important role in the signal processes that determine whether a cell will undergo cellular division, cellular differentiation, or cellular death. These findings have important implications on the ability to regulate cell division as well as sensitivity and resistance to chemotherapeutic agents and, therefore, for treating diseases associated with excessive cellular division, aberrant differentiation, and premature cellular death, e.g., for the treatment of cancers, autoimmune disease, degenerative diseases, regeneration, etc.
Mitochondrial uncoupling proteins (UCP 1-5) are a family of molecules, first described in brown adipose tissue, that. dissipate of the proton gradient, lower mitochondrial membrane potential, induce of a metabolic shift to fatty acids as a source of fuel in mitochondria, promote of high rates of glucose utilization in the cytosol and increased oxygen consumption in the mitochondria; and reduce production of reactive oxygen species [29,30]. UCP-mediated uncoupling requires transport of the anionic fatty acids head groups by UCP and proton transport via the bilayer [31]. UCP-2 has been shown to be expressed at higher levels in the mitochondria of cells that are protected from apoptosis-inducing stimuli. [32]. Data is consistent with the report that the production of reactive oxygen species and subsequent cellular damage induced by hyperglycemia in endothelial cells is prevented by transfection with UCP-1. UCP are inducible proteins. For example, it has been shown that lipopolysaccharides can induce synthesis of UCP-2 [33] and superoxide can activate UCP-2 within the mitochondria [34]. UCP-2 mRNA and protein are induced in the cerebral cortex after trauma {Bechmann, 2002 #81} [35] ischemic preconditioning in vivo or intraperitoneal injection of kainic acid [36,37], and UCP-4 is inducible in neurons in vitro by glycolytic inhibitors [38]. Significant neuroprotective actions of UCP-induced uncoupling have been shown in vitro models of oxygen-glucose deprivation [39] and in vivo [39] models of trauma ischemia, seizures [39] and excitotoxicity [37] in mice overexpressing UCP.
Several cell surface proteins have previously been identified as cell death proteins. These proteins are believed to be involved in initiating a signal which instructs the cell to die. Cell death proteins include, for example, Fas/CD95 (Trauth, et al., Science, 245:301, 1989), tumor necrosis factor receptors, immune cell receptors such as CD40, OX40, CD27 and 4-1BB (Smith, et al., Cell, 76:959, 1994), and RIP (U.S. Pat. No. 5,674,734). These proteins are believed to be important mediators of cell death. These mediators, however, do not always instruct a cell to die. In some cases, these mediators actually instruct a cell to undergo cell division. The intracellular environment, and particularly the status of the proton motor force and the source of fuel for mitochondrial metabolism, determines whether stimulation of the cell death protein will lead to a signal for death or cell division (see, e.g., U.S. patent application Ser. No. 09/277,575, incorporated herein by reference).
These findings have important implication in the present invention in dealing with drug resistant tumors. Every year at least 6.2 million people die worldwide from cancer [1]. Many cancer patients will be treated by chemotherapy. For some people this treatment will be effective, but it many cases chemotherapy is not successful, in part because of the development of drug resistance. It is commonly observed in treating cancers, that initial treatments, such as with chemotherapy and/or radiation therapy, are effective to destroy significant numbers of tumor cells, only to leave behind a small number of tumor cells that are resistant to the treatment, which then multiply to form newly detected tumors that are increasingly resistant to treatment as new rounds of therapy are tried. The growing popularity of “cocktails” of chemotherapy drugs has given rise to multidrug resistant (“MDR”) tumor cells, which are ever more difficult to destroy. Drug sensitive tumor cells, under the selective pressure of treatment with drugs, develop into drug resistant versions of the same tumor cell type. It is the drug resistant cells that take over, and with each round of chemotherapy the proportion of drug resistant cells to drug sensitive cells increases, to the point where recovery becomes more and more difficult, and eventually the cancer becomes untreatable. Indeed, drug resistance, either acquired or inherent, is the leading cause of death in cancer [27]. Mechanisms which have been suggested to account for drug resistance include over-expression of a multi-drug resistance transporter (pgp-1) [27], failure to express death inducing receptors [27, 28], and a metabolic strategy that may provide protection from a variety of stresses. Because drug resistance is such an important problem, one of the goals of the present invention is to provide methods to overcome this problem. Methods for dealing with MDR tumor cells have been proposed, but without practical, clear clinical success at entirely eliminating such cells and providing a cure for patients with MDR tumors. For example, in Lampdis and Priebe U.S. Pat. No. 6,670,330, entitled: “Cancer Chemotherapy with 2-Deoxy-D-Glucose”, incorporated herein in its entirety by reference, a class of glycolytic inhibitors are described for use in combination with standard chemotherapy protocols in treating solid tumors by attacking anaerobic cells a the center of the tumor.